Homeostasis of continuously renewing tissues, such as the epidermis, is maintained by somatic stem cells. These are undifferentiated, self-renewing cells, which also produce daughter transient amplifying (TA) cells that make up the majority of the proliferative population. TA cells undergo a finite number of cell divisions before leaving the proliferative compartment and moving toward terminal differentiation, whereas it has been assumed that the stem cells persist throughout the lifetime of the organism. Through a series of labeling experiments with tritiated thymidine, we previously showed that stem cells from adult mouse skin did not divide as often as the other basal cells, but they did divide at a steady rate in vivo. We also showed that they continued to proliferate in vivo throughout life, and that they have a high proliferative potential in vitro. Thus, it may be that epidermal stem cells do not follow the Hayflick theory and possibly "live forever." Last year, we developed a sorting method that yields a viable population of stem cells from the epidermis. We showed that these stem cells have the capacity to regenerate the epidermis and to continuously express a recombinant gene in the regenerated tissue. Thus, they are the stem cells for the epidermis. Very recently, we determined that these epidermal stem cells also have the remarkable ability to participate in the formation of the other tissues, a plasticity similar to that of embryonic stem cells and a few other somatic stem cells. Our preliminary data show that the epidermal stem cells isolated from neonatal mouse skin incorporate into a variety of tissues and alter their phenotype after injection into blastocysts. Thus, the fate determination of these stem cells appears to be in direct response to their environment. We also determined that the cell cycle profile of these cells appears to be an intrinsic property of the stem cell and not related to the age of the cell. These data have led us to ask what happens to epidermal stem cells as the organism ages? In this application, we propose to test whether the age of the stem cells has an effect on its response to the environment or extrinsic factors in determining its fate pathway. We propose the following specific aims: 1) to compare old vs. young murine epidermal stem cells by determining their number, altered phenotype and longevity after injection into blastocysts, and to determine whether there is a difference in the life span of mice derived from blastocysts injected with old vs. young stem cells; and 2) to compare old vs. young adult human epidermal stem cells by determining if there is a difference in their in vitro growth potential and if they respond differently to extrinsic environmental signals in vitro and after injection into blastocysts.